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Title: Mapping transmembrane binding partners for E-cadherin ectodomains

We combine proximity labeling and single molecule binding assays to discover transmembrane protein interactions in cells. We first screen for candidate binding partners by tagging the extracellular and cytoplasmic regions of a “bait” protein with BioID biotin ligase and identify proximal proteins that are biotin tagged on both their extracellular and intracellular regions. We then test direct binding interactions between proximal proteins and the bait, using single molecule atomic force microscope binding assays. Using this approach, we identify binding partners for the extracellular region of E-cadherin, an essential cell–cell adhesion protein. We show that the desmosomal proteins desmoglein-2 and desmocollin-3, the focal adhesion protein integrin-α2β1, the receptor tyrosine kinase ligand ephrin-B1, and the classical cadherin P-cadherin, all directly interact with E-cadherin ectodomains. Our data shows that combining extracellular and cytoplasmic proximal tagging with a biophysical binding assay increases the precision with which transmembrane ectodomain interactors can be identified.

Authors:
; ; ;
Publication Date:
NSF-PAR ID:
10202824
Journal Name:
Proceedings of the National Academy of Sciences
Volume:
117
Issue:
49
Page Range or eLocation-ID:
p. 31157-31165
ISSN:
0027-8424
Publisher:
Proceedings of the National Academy of Sciences
Sponsoring Org:
National Science Foundation
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